This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Glucose stimulates both insulin secretion and hydrolysis of arachidonic acid (AA) esterified in membrane phospholipids of pancreatic islet [unreadable]-cells, and these processes are amplified by muscarinic agonists. Here we demonstrate that nonesterified AA regulates the biophysical activity of the pancreatic islet [unreadable]-cell delayed rectifier channel, Kv2.1. Recordings of Kv2.1 currents from INS-1 insulinoma cells incubated with AA (5 [unreadable]M) and subjected to graded degrees of depolarization exhibit a significantly shorter time-to-peak current interval than do control cells. AA causes a rapid decay and reduced peak conductance of delayed rectifier currents from INS-1 cells and from primary [unreadable]-cells isolated from mouse, rat, and human pancreatic islets. Stimulating mouse islets with AA results in a significant increase in the frequency of glucose-induced [Ca2+] oscillations, which is an expected effect of Kv2.1 channel blockade.. Stimulation with concentrations of glucose and carbachol that accelerate hydrolysis of endogenous AA from islet phosphoplipids also results in accelerated Kv2.1 inactivation and a shorter time-to-peak current interval. Group VIA Phospholipase A2 (iPLA2[unreadable]) hydrolyzes [unreadable]-cell membrane phospholipids to release nonesterified fatty acids, including AA, and inhibiting iPLA2[unreadable] prevents the muscarinic agonist-induced accelerated Kv2.1 inactivation. Stably transfected INS-1 cells that overexpress iPLA2[unreadable] hydrolyze phospholipids more rapidly than control INS-1 cells and also exhibit an increase in the inactivation rate of the delayed rectifier currents. ESI/MS/MS analyses of the phospholipid composition of the genetically manipulated insulinoma cells revealed neither deficiency or augmentation of their ability to synthesize arachidonate-containing species. These results suggest that Kv2.1 currents could be dynamically modulated in the pancreatic islet [unreadable]-cell by phospholipase-catalyzed hydrolysis of membrane phospholipids to yield non-esterified fatty acids, such as AA, that facilitate Ca2+ entry and insulin secretion.